Abstract: The overall goal of Skyran Biologics is to develop a powerful topical therapeutic to combat diabetic retinopathy (DR), a leading cause of vision loss and total blindness globally. Our research team was one of the first to show that a novel protein, Pigment Epithelium-Derived Factor (PEDF) is expressed in the eye and provides significant protection to the injured retina. After identifying a 44-mer bioactive region of PEDF, we generated several structurally different analogs to this region, screened these for bioactivity, and identified Spx81-5 as a lead compound with improved activity in the retina over the native fragment. These proof of concept studies showed that topical Spx81-5 can safely and effectively restore multiple markers of DR to near control levels when applied topically to the Ins2Akita mouse model of DR. In both the mouse model and preliminary experiments on non-human primates, we have shown that topical Spx81-5 leads to therapeutic concentrations of the peptide in the vitreous. We will use two specific aims to address three major scientific issues: The first is to confirm the proof of concept for Spx81-5 and to show that its efficacy is specific to neither species nor mechanism of inducing DR. The second is to confirm that Spx81-5 improves visual acuity. The third is to verify that Spx81-5 is available in the vitreous and/or retina via drop form in eyes where there is substantial back-to-front vitreal flow. Skyran Biologics Inc. can significantly increase the value of Spx81-5 by definitively demonstrating that the drop formulation of Spx81-5 results in therapeutic doses of the peptide in the vitreous and retina. Our first aim in this project is to validate efficacy of Spx81-5 using pharmacologically (STZ) induced diabetes in Long-Evans rats. This model has been widely validated in preclinical DR studies and used to show proof of concept for improvement in vision, through visual acuity and contrast sensitivity measurements. We will also assess efficacy of Spx81-5 to reduce retinal cell death and inflammation. In Aim 2, we will analyze biodistribution of Spx81-5 when given topically to adult rabbits to show that topical administration will be effective as a route of delivery. We will deliver radioactively-labeled Spx81-5 to the eye topically at two different doses and measure distribution of labeled peptide in aqueous, vitreous and retina to confirm that the data already demonstrated using drop form can be replicated in a larger eye. The milestones achieved by the successful completion of these aims will position us to propose a set of key Phase II studies including systemic toxicity screening, dosing, formulation, biostability and the development of GMP grade peptide for clinical translation. The Phase II work will move us towards essential IND-enabling studies for FDA approval and development of clinical phase translational strategies.